Thromboembolic vascular disease comprises a significant fraction of all morbidity and mortality in the United States. Thrombolytic therapy, using systemically or selectively administered plasminogen activators, has gained increasing acceptance for the treatment of thromboembolic disease. However, further improvements in thrombolytic therapy are still required.
Various compounds have been employed for the intravascular dissolution of thrombi and emboli in mammals. For example, heparin, coumarins, trypsin, chymotrypsin, streptokinase, urokinase, staphylokinase, plasmin and certain snake venoms have been studied and shown to be fibrinolytic in vitro. However, these fibrinolytic agents each produce certain undesirable side effects. For example, anti-coagulants such as heparin, while useful in many instances, must be used with extreme caution in disease states where there is increased danger of hemorrhage.
Urokinase and streptokinase operate by the conversion of plasminogen to the proteolytic enzyme plasmin, which degrades fibrin clots as well as other plasma proteins. When plasmin formed by these plasminogen activators exceeds the capacity of the circulating plasmin inhibitor, .alpha.2-antiplasmin, fibrinogen and several other clotting factors will be depleted, enhancing the probability of hemorrhagic complications, common after thrombolytic therapy. Plasminogen activators also consume plasminogen and may deplete this protein to a level at which further thrombolysis is prohibited. Thus, it has been a desideratum to provide a fibrinolytic agent which acts directly on the appropriate thrombus or embolus, without causing a systemic lysis of fibrin deposits and other plasma proteins.
It is known that various snake venoms actively lyse human blood clots and may have certain advantages over more common fibrinolytic agents. However, such venoms have been shown to include fibrinogenolytic, thrombic, thromboplastic, hemolytic or hemorrhagic properties which contra-indicate their clinical use. While fractionation of the whole venoms has been attempted, the isolation of a pure fibrinolytic fraction having clinical usefulness and which is devoid of toxic activities has not been attained.
According to the present invention, a metalloproteinase moiety having direct fibrinolytic action is provided and isolated from snake venom. The moiety has a molecular weight of from about 25 to 27 kilodaltons (kd) and an isoelectric point of from about 6.5 to 7.0, is free of the toxic side effects hereinbefore described and has a direct fibrinolytic activity that is not readily inhibited by serum antiprotease when employed in vivo. The pure venom fraction may be isolated by the novel combinations of diverse fractionations hereinafter described, or the moiety or active portions thereof may be produced by biochemical methods such as genetic engineering or the like. Thrombolytic therapy is now established as a procedure for the treatment of acute deep-vein thrombosis and pulmonary embolism, and the purified enzyme of the present invention presents a substantial therapeutic advantage as it is able to rapidly lyse thrombi with minimal systemic toxicity.